Researchers at the U.S. Department of Energy's Argonne National Laboratory and the University of Chicago have determined the crystal structure of sortase B, an enzyme found in the bacteria that cause staph and anthrax. While an antibiotic is probably five to seven years away, the structure could provide the first clue in developing a treatment for the infections.
The research is published today in the journal Structure.
It took the researchers 21 days to build the three dimensional model of sortase from the genome. Without the new technology available at Argonne's Structural Biology Center, including the Advanced Photon Source's powerful X-rays to illuminate the structures and the Midwest Center for Structural Genomics' robotic and automation facilities for protein expression, purification and crystallization, the process could have taken several months.
By analyzing genomes, the researchers uncover information that will lead to structure-based or "rational" drug design. The problem is that researchers don't know what half the proteins coded by the genome do or how they work.
Now that the researchers understand the enzyme, they hope to find a way to stop it – or at least to slow it down. Sortase attaches proteins to the surface of bacterial pathogens. These proteins help the pathogens survive and flourish.
Bacteria like staph and anthrax need iron to function. But little free iron is available in the blood stream because most of it is bound in red blood cells. So the bacteria develop a mechanism to pry open the red blood cells, and these proteins help them.
"This is actually a very smart mechanism," said Andreji Joachimiak, lead researcher and director of the Structural Biology Center. The process is outlined in an article published in Science last year by Olaf Schneewind of the University of Chicago, which laid the groundwork for the sortase project.
The bacteria open the blood cell, bind the hemoglobin that contains heme – the pigment containing iron in hemoglobin – transport the heme, degrade the heme and then extract the iron.
Before the protein can bind the hemoglobin, it has to be attached to a specific position on the surface of the cell. The bacteria use a specific enzyme to accomplish this; in this case it is sortase.
"Sortase would be a good target for a drug, because if one can block the enzyme, it will not be able to attach these proteins to the surface and the bacteria would not be able to get iron from our bloodstream," Joachimiak said.
The research looks at sortase from both staph and anthrax – more formally, Staphylococcus aureus, and Bacillus anthracis, and concludes that the two are similar. Both have the same catalytic amino acid triad with Cys, His and Asp residues present in both enzymes -- which means that the site of the enzyme-protein reaction is the same. Only the location of one of the residues varies.
Joachimiak said the fact that they have the same triad is important. If the sortase active site is the same in both, it can be blocked with just one drug. Furthermore, versions of sortase are found in several other gram positive bacteria. That means one drug could double up and target a variety of different bacteria. Also significant, the enzyme is found only in gram positive bacteria, meaning treatments that target it would not likely affect human enzymes.
Now that the structure is known, Joachimiak said the next step is to mimic the signal sequence, or peptide, in the protein with a drug that blocks the enzyme.
"We would like to design a drug that will look like the peptide, but will not be the peptide," he said. "Something else that will bind to the same site and make sure the enzyme is dead or inactive."
This step is based primarily on trial and error. However, if scientists know the structure, they can make a more educated guess. "We need to study more proteins from these genomes to better understand their biology and therefore be able to treat them or control them," Joachimiak said. "We know so little so far."
Research continues at Argonne's Structural Biology Center where more than 530 structures have been determined. Nearly 150 protein structures have been determined at the Midwest Center for Structural Genomics and recorded with the International Protein Data Bank -- that's more than any other structural genomics center.
Joachimiak's co-authors are colleagues R-g. Zhang, R-y. Wu and G. Joachimiak at Argonne and S.K. Mazmanian, D.M. Missiakas, P. Gornicki and O Schneewind from the University of Chicago. The published research was supported by the National Institutes of Health Grants and the U.S. Department of Energy Office of Biological and Environmental Research.
The nation's first national laboratory, Argonne National Laboratory conducts basic and applied scientific research across a wide spectrum of disciplines, ranging from high-energy physics to climatology and biotechnology. Argonne has worked with more than 600 companies and numerous federal agencies and other organizations to help advance America's scientific leadership and prepare the nation for the future. The University of Chicago operates Argonne as part of the U.S. Department of Energy's national laboratory system.
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